Chemistry: molecular biology and microbiology – Micro-organism – per se ; compositions thereof; proces of... – Bacteria or actinomycetales; media therefor
Reexamination Certificate
2001-02-05
2003-04-08
Lilling, Herbert J. (Department: 1651)
Chemistry: molecular biology and microbiology
Micro-organism, per se ; compositions thereof; proces of...
Bacteria or actinomycetales; media therefor
C435S262500, C588S253000
Reexamination Certificate
active
06544773
ABSTRACT:
FILED OF THE INVENTION
The invention relates A novel marine microorganism (Pseudomonas CH07) isolated from the Indian coastal zone near Goa which is capable of biodegradation of PCBs including sterically hindered di and tri-ortho chlorinated biphenyls and coplanar congeners present in a technical grade Clophen A-50 (Bayer, Lot no. 16572) by the novel strain of marine microorganism, Pseudomonas CH07.
BACKGROUND OF THE INVENTION
Classification of PCBs
PCBs are non-volatile organic compounds (depending on the degree of chlorination they may belong to the class of semi volatile organic compounds (SVOC). PCBs are divided into two groups of isomers based on the difference in structure:
1. Coplanar PCBs—those compounds have chlorinated substituents in both para positions, and any/all meta positions. Meta or para chlorine substituents have, by their structure, low steric hindrance with neighboring H, which allows free rotation about the phenyl-phenyl bond. There are 20 coplanar PCBs, out of which three (77, 126, 105) are very toxic. Most importantly, they are non-ortho chlorinated.
2. Mono-ortho chlorinated PCBs-all molecules have one-chloro substitutions in the other positions only. Ortho substituents tend to create rigid bonds due to the large steric interference between C1 and H atoms.
3. Di-ortho a represents the chlorine atoms at the ortho-positions.
4. Tri-ortho represents the chlorine atom at the ortho-positions.
Depending on the number of C1 atoms on the biphenyl rings PCBs are divided into mono, di, tri, tetra, penta, hexa, hepta, octa, nona and deca-chlorobiphenyls and there are 209 possible PCBs theoretically but many do not occur because of steric hindrance.
Most abundant PCBs in commercial mixtures are ortho-substituted congeners, which are readily degradable. However, smaller amounts of the so-called ‘di-oxin’ like PCBs namely the coplanar (=non-ortho substituted) and mono-ortho substituted congeners, are present in the commercial mixture as well. Broad classification:
a) Non-ortho or mono-ortho chlorinated Biphenyls (coplanar).
b) Ortho-substituted chlorinated biphenyls.
c) Sterically hindered chlorinated Biphenyls.
Polychlorinated biphenyls (PCBs) represent a class of toxic xenobiotics that are distributed throughout the biosphere. Over the past several years, PCBs have received increasing attention due to accumulation of their residues in tissues of living organisms and biomagnified through the food chain leading to health hazards. PCBs are produced by direct chlorination of biphenyl. Due to the large number of hydrogen atoms present on the biphenyl nucleus, many different chlorinated compounds (termed “congeners” isomers of different homologous series) are possible. As many as 209 congeners of the PCBs could be theoretically produced [Furukawa, Biodegradtion and Detoxification of Environmental Pollutants, p. 34-57. CRC press. (1982)]; however, due to steric restrictions, only about half of this number are actually found in the environment. Therefore, PCBs are mixtures of a variety of chlorine-substituted biphenyl molecules. Clophen A-50 is a technical grade chemical compound containing about 40 different congeners of PCBs. Clophen is well known in commercial circles and to peoples skilled in the art. In fact, Clophen A-50 is equivalent to Aroclor 1256 i.e. it has about 56% chlorines w/w [Yadav; Jagjit S.; Reddy; C. A.; Quensen; John F.; Tiedje; James M. Degradation of polychlorinated biphenyl mixtures in soil using
Phanerochaete chrysosporium
in nutrient rich, non-ligninolytic conditions. U.S. Pat. No. 6,107,079. (Aug. 22, 2000)].
Due to their lipophilic and hydrophobic characteristics, the PCBs get accumulated in tissues of various species of organisms and are magnified through the food chain [Furukawa supra; Jacobson et al. Develop. Psychol. 20: p.523-532. (1984); Sarkar, A. and Everaarts, J. M. (1998) Riverine input of chlorinated hydrocarbons in the coastal Pollution. In: Ecology of Wetlands and Associated Systems. Ed. S. K. Majumdar, E. W. Miller and Fred J. Brenner. Chapter 27, Pub: Pennsylvania Academy of Science. pp, 400-423; Sarkar, A. (1994) Occurrence and distribution of persistent chlorinated hydrocarbons in the seas around India. In: The Oceans: Physico-chemical Dynamics and Resources (ed) S. K. Majumdar, E. W. Miller, G. S. Forbes, R. F. Schmalz and Assad, A. Panah. The Pennsylvania Academy of Science. Chapter-28,pp, 445-459.]. The physical effects of PCBs vary from mammals, to birds, to humans. Natural microbial populations do not easily remove PCBs. Some PCB congeners are found to be transformed by both anaerobic and aerobic bacteria [Abramowicz, D. A., Crit. Rev. Biotechnol. 10: 241-251. (1990)]. The aerobic degradation of PCBs is generally limited to less—chlorinated congeners (five or fewer chlorines per biphenyl molecule) by an enzymatic mechanism involving deoxygenation of the aromatic ring [Bedard, D. L., et al., Appl. Environ. Microbiol. 53: 1094-1102. (1987); Bradley; Clifford A.; Kearns; Robert D.; Wood; Pauline P.; Black; William E. Degradation of polyhalogenated biphenyl compounds with white-rot fungus grown on sugar beet pulp. U.S. Pat. No. 5,583,041 (Dec. 10, 1996); Sarkar, A. (1994) Comments on “Degradation of polychlorinated dibenzo-p-dioxin and dibenzo-furan contaminants in 2,4,5-T by photoassisted iron-catalyzed hydrogen peroxide” by J. J. Pignatello and L. Q. Huang. Wat. Res. Vol. 27: 1731-1736. Water Research. Vol: 28 No. 12, pp, 2589-2594. Sarkar, A. (1994) Comments on: “Evaluation of dechlorination mechanisms during anaeorobic fermentation of bleached kraft mill effluent”, by W. J. Parker, E. R. Hall and G. J. Farquhar, Wat. Res. 27, 1269-1273 (1973). Water Research Vol. 28: No. 9, pp, 2043-2044, 1994.] The more chlorinated congeners are generally recalcitrant to aerobic degradation [Kimbara; Kazuhide; Shimura; Minoru; Hatta; Takahasi; Kiyohara; Hohzoh. Method for degrading polychlorinated biphenyls and novel microorganism. U.S. Pat. No. 5,897,996 (Apr. 27, 1999)]. In 1978, Furukawa and associates [Furukawa et al., Appl.Environ. Microbiol. 35:223-227. (1978)] studied the biodegradability of several isomers of PCBs. They found that as chlorine substitution increased, degradability decreased. An isomer with four Cl was not easily degraded. The position of the chlorine is also important. Ortho positioning of two chlorines on a single ring greatly inhibited degradation [MacFaddin, F. J. 1980. Biochemical tests for identification of medical bacteria. Second edition. p. 527. Williams and Wilkins, Baltimore].
Considering the environmental importance of PCBs and the hazards posed by them, numerous investigators have been examining biological detoxification systems to deal with PCBs. One way to decipher the complexities of highly chlorinated isomers is to look into the problem, according to the order of the extent of chlorine substitution in the biphenyl ring. Of the 209 theoretically possible isomers and congeners of PCB, 20 members attain coplanarity due to non-ortho chlorine substitution in the biphenyl rings. In this group, three coplanar congeners such as 3,3,4,4′-tetrachlorobiphenyl, 3,3′,4,4′,5-pentachlorobiphenyl and 3,3′,4,4′-pentachlorobiphenyls and 3,3′,4,4′,5,5′-hexachlorobiphenyls are approximate isostereomers of highly toxic 2,3,7,8-tetrachlorodibenzo-p-dioxin and 2,3,4,7,8-pentachlorodibenzofuran and hence elicit similar toxic and biologic responses typical of dioxins and furans [Safe, S.(1984) Polychlorinated biphenyls (PCBs) and polybrominated biphenyls (PBBs): biochemistry, Toxicology and mechanism of action. CRC Crit. Rev. Toxicol., 13, 319-93].
Because of the coplanarity, these three congeners of PCB were found to be most toxic chlorinated aromatic compounds in comparison with the rest of the congeners of PCBs [Yoshihara, S.; Nagata, K.; Yoshimura, H.; Kuroki, H. and Masuda, Y. (1981). Inductive effect on hepatic enzymes and acute toxicity of individual polychlorinated
De Jaysankar
Nagappa Ramaiah
Sarkar Anupam
Council of Scientific & Industrial Research
Greenspan Myron
Lackenbach Siegel
Lilling Herbert J.
Nissen J. Harold
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